Project Summary/Abstract Colon cancer and colorectal liver metastases (CRLM) are a significant and increasingly lethal disease. Though there is a strong scientific premise to harness the immune system to treat this cancer, there remains a funda- mental gap in understanding of how immunotherapies can be utilized for the majority of these tumors, particu- larly microsatellite stable cancers. The central hypothesis is that the tumor microenvironment can be manipu- lated to enhance cytotoxic lymphocyte infiltration and activation, and that checkpoint blockade can be simulta- neously utilized to incite a clinically relevant immune response. This ?one-two punch? hypothesis has been formulated on the basis of preliminary data produced by the applicant where increased T-cell trafficking to tu- mors, when combined with CTLA4 blockade, resulted in complete CRLM regressions. The rationale for the proposed research is that once it is known how to enhance immunostimulatory signals in the microenvironment and simultaneously suppress inhibitory influences, a new strategy for the management of colon cancer is pos- sible. Supported by a very strong scientific premise based on published papers and robust preliminary data, this hypothesis will be tested by pursuing three specific aims: 1) Determine mechanisms to enhance lympho- cyte proliferation and anti-tumor specific immune responses in colon cancer by manipulating immunosuppres- sive signals, 2) Examine mechanisms that supply immunostimulatory influences directly into the tumor; and 3) Combine checkpoint blockade with selective delivery of human LIGHT to treat surgically resected tumors and human CRLM in a pre-clinical autologous system. Under the first aim we expect to increase lymphocyte infiltra- tion, proliferation and activation while simultaneously curbing immunosuppressive signals/cells in the microen- vironment utilizing a validated pre-clinical model established by the applicants. In the second aim, a clinically relevant method to safely increase LIGHT expression within colon cancer tumors using novel tumor-specific viral delivery mechanisms engineered by the applicants will be analyzed. Under the third aim, patient tumors will be utilized in an autologous humanized mouse model and treated with CTLA4 blockade combined with on- colytic viral delivery mechanisms to increase LIGHT expression. The proposed research is innovative, because the multi-combination therapy of LIGHT expression in CRLM with tumor-specific oncolysis and checkpoint blockade will deliver an inventive approach that will be universally applicable from patient to patient. New hori- zons that will stem from this innovative strategy include a better understanding of anti-CTLA4 biology that may not only enhance response rates, but also vastly increase indications for its use in previously ?cold? tumors, including microsatellite stable gastrointestinal cancer. This contribution will be significant because it will estab- lish a synergistic combination of immunotherapies that will have direct translational impact on one of the dead- liest cancers worldwide. The implications of our results may improve patient quality of life and provide survival advantages over the best current surgical and chemotherapeutic strategies for this disease.